Radiolabelling of Polyclonally Expanded Human Regulatory T Cells (Treg) with 89Zr-oxine for Medium-Term In Vivo Cell Tracking

Abstract

Regulatory T cells (Tregs) are a promising candidate cell therapy to treat autoimmune diseases and aid the longevity of transplanted solid organs. Despite increasing numbers of clinical trials using human Treg therapy, important questions pertaining to their in vivo fate, distribution, and function remain unanswered. Treg accumulation in relevant tissues was found to be crucial for Treg therapy efficacy, but existing blood-borne biomarkers are unlikely to accurately reflect the tissue state. Non-invasive Treg tracking by whole-body imaging is a promising alternative and can be achieved by direct radiolabelling of Tregs and following the radiolabelled cells with positron emission tomography (PET). Our goal was to evaluate the radiolabelling of polyclonal Tregs with 89Zr to permit their in vivo tracking by PET/CT for longer than one week with current preclinical PET instrumentation. We used [89Zr]Zr(oxinate)4 as the cell-labelling agent and achieved successful radiolabelling efficiency of human Tregs spanning 0.1–11.1 Bq 89Zr/Treg cell, which would be compatible with PET tracking beyond one week. We characterized the 89Zr-Tregs, assessing their phenotypes, and found that they were not tolerating these intracellular 89Zr amounts, as they failed to survive or expand in a 89Zr-dose-dependent manner. Even at 0.1 Bq 89Zr per Treg cell, while 89Zr-Tregs remained functional as determined by a five-day-long effector T cell suppression assay, they failed to expand beyond day 3 in vitro. Moreover, PET imaging revealed signs of 89Zr-Treg death after adoptive transfer in vivo. In summary, 89Zr labelling of Tregs at intracellular radioisotope amounts compatible with cell tracking over several weeks did not achieve the desired outcomes, as 89Zr-Tregs failed to expand and survive. Consequently, we conclude that indirect Treg labelling is likely to be the most effective alternative method to satisfy the requirements of this cell tracking scenario.

Labeling Natural Killer cells with superparamagnetic iron oxide nanoparticles for detection by preclinical and clinical-scale magnetic particle imaging

“…57 Unfortunately, nuclear imaging will not fulfill our clinical need to track delicate NK cells in pediatric populations and due to potential cytotoxic effects of 89 Zr on cell activity even at low doses per cell. 30 As we demonstrate in this work, MPI to addresses the limitations of current cellular imaging techniques. MPI specifically identifies the location of iron-labeled cells and produces a specific (hotspot) image [31][32][33][34][35][36][37][38] , similar to images obtained by 19 F MRI and nuclear imaging.…”

“…57 Unfortunately, nuclear imaging will not fulfill our clinical need to track delicate NK cells in pediatric populations and due to potential cytotoxic effects of 89 Zr on cell activity even at low doses per cell. 30…”

“…[27][28][29] Nuclear imaging techniques (PET and SPECT) produce excellent image contrast that is quantitative but can be detrimental to cell viability. 30 Magnetic particle imaging (MPI) is an emerging modality for in vivo tracking of fate, biodistribution, quantification, and persistence of cellular therapeutics. To enable cell detection by MPI, cells are prelabeled in culture with iron oxide nanoparticles.…”

Biomaterial-enhanced treg cell immunotherapy: A promising approach for transplant medicine and autoimmune disease treatment

“…In a study by Jacob et al, polyclonal expanded human Tregs were successfully tracked and monitored using radiolabeled 89 Zr-oxine in immunodeficient mice for three days. The results showed that this radiolabeling did not affect the viability of Tregs or their suppressive activities [ 147 ]. Another study involved radiolabeling of CD4 + T cells with Tc-99 m (Technetium-99 m pertechnetate), enabling Treg biodistribution assessment within a day [ 149 , 154 ].…”